Electrochemical analysis of formation of polynucleotide complexes in solution and at electrode surfaces

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The formation of double- and triple-stranded complexes between the synthetic homopolyribonucleotides in solution and at the surfaces of the hanging mercury drop (HMDE) and carbon paste (CPE) electrodes was studied by means of voltammetric and chronopotentiometric methods. It was shown that the cyclic voltammetry reduction signal of poly(C) and the anodic signal of poly(G) obtained with HMDE, as well as the chronopotentiometric oxidation signal of poly(G) obtained with CPE can be used to detect the duplex and triplex formation in solution. In addition to these intrinsic polynucleotide signals, the indicator [Co(phen) 3] 3+ was used to differentiate between single-stranded and ordered structures at CPE. It was shown that the indicator binds preferentially to the duplex poly(G) · poly(C) (formed in solution followed by adsorption at CPE) displaying, however, no preferential binding to the triplex structures. If poly(C) or poly(G) were immobilized at the electrode surface (by adsorption forces) and hybridized with the complementary polynucleotide in solution (at neutral pH), only a small extent of complexation could be detected at HMDE under the given conditions. On the other hand, analogous experiments with CPE suggested formation of ordered polynucleotide structures at the carbon electrode surface. As a result of complexation of poly(C) with poly(G) at neutral pH, formation of more than one ordered structure, including the duplex, was indicated. Polynucleotide complexation performed at pH 3.3 (optimum for formation of the protonated triplex poly(C +) · poly(G) · poly(C) in solution) resulted in the formation of a mixture of structures including triplex and duplex structures. Binding of [Co(phen) 3] 3+ to the product of poly(A) complexation with poly(U) suggested that the yield of the duplex formed at the CPE surface may be influenced by the conformation of the single-stranded polynucleotide anchored at the surface prior to the hybridization. The implications of these findings to DNA biosensor development are discussed.